We have shown that the activity of heme synthetase, which catalyzes the chelation of iron to protoporphyrin to form heme, is deficient in cultured skin fibroblasts from patients with protoporphyria. This inherited disorder is characterized by excessive protoporphyrin accumulation and excretion. Using fibroblasts as the source of heme synthetase, we shall determine if protoporphyric cells contain an enzyme species with a different Michaelis constant for porphyrin and/or iron. The enzyme will be solubilized in order to study some of its properties, in particular that it may be a dimer. The dominant effect of the abnormal gene will be examined by cell hybridization, and the chromosomal location of the genes coding for synthesis of heme synthetase and other heme pathway enzymes will be studied with panels of rodent-human cell hybrids. Cultured fibroblasts will also be used to investigate other aspects of abnormal protoporphyrin metabolism. The possibility that heme synthetase and/or protoporphyrinogen oxidase is deficient in variegate porphyria will be assessed. In bovine protoporphyria, heme synthetase activity in cultured fibroblasts from affected cows will be compared with that in their parents and in normal cows to examine the hypothesis that this is a recessively inherited disease. The residual enzymatic activity will be characterized as in human protoporphyria. The reson why the zinc chelate of protoporphyrin accumulates in lead poisoning and iron deficiency, and not free protoporphyrin, will be examined by incubating fibroblasts in monolayer culture with ALA and variable combinations of iron, zinc, and lead. The amounts of free protoporphyrin and zinc protoporphyrin which are formed under the various conditions will be determined. The cell system will also be used to study the mechanism by which protoporphyrin causes photo-induced tissue damage, and the protective effect of B-carotene and other compounds will be examined.